臺灣博碩士論文加值系統

血吸蟲病的危害遍及全球，其中曼生血吸蟲主要分布於非洲、美洲及加勒比海群島。目前已從蟲體內找出許多可供疫苗發展的蛋白分子，sm22.6即為其一。許多研究都指出重組蛋白sm22.6可引發免疫球蛋白E型 (IgE) 的辨識，並且病人體內的IgE含量和蟲體的再感染的程度更被證實具有負相關性，因此對其寄予製成疫苗的期望。不幸的是，往後的動物接種實驗卻發現單就此抗原所引發的抵抗力不足抵抗蟲體再感染，加重了疫苗研發的困難。由於在過去的研究中對sm22.6抗原的生物化學及生物物理特性缺乏了解，因此希望能從生物化學及細胞生理學兩方面去探討此抗原的分子特徵。目前，此重組蛋白sm22.6已能成功地在大腸桿菌(BL21DE3)裡表達且純化出相當純度的rsm22.6。經由變性及非變性蛋白電泳的生化分析，顯示出sm22.6有著高度的疏水性，並且導致此抗原嚴重的多聚化。雖亦使用了其他的測試方法，包括核磁共振及DNA結合測試，但皆不足以提供進一步的結果。於是決定利用人類乳癌細胞 (MDA-MB-435s) 建立sm22.6的真核表達系統並定位此抗原的細胞位置。結果發現sm22.6散佈於整個細胞內部，與其在大腸桿菌中表達出的蛋白分子特性並不相似，而其生理功能似乎亦不為先前利用蛋白質體學的研究所推測的細胞骨架。

Schistosomiasis is one of the most widespread parasite diseases in the world, whereas Schistosoma mansoni is a major schistosome species in Africa, America, and the Caribbean islets. Many antigenic vaccine candidates have been postulated, including sm22.6 and GST. Although the lower level of re-infection of human schistosomiasis is related to the higher level of IgE against rsm22.6, unfortunately, the observation of the experimental vaccination in mice finds some difficulties in further development of vaccine. In addition, the biochemical and biophysical properties of the antigen are virtually unknown, thus the present study intends to characterize sm22.6 from biochemistry and cell biology. To do this, sm22.6 was expressed in E. coli (BL21DE3) and purified to homogeneity. Analyses of the recombinant protein showed that the antigen was highly hydrophobic and formed polymers readily as judged by both native and denatured electrophoreses. Because various technologies including NMR and DNA binding which had been applied to the study of the antigen generated vague results, we decided to express the antigen in human breast cancer cell (MDA-MB-435s) to locate in the subcellular compartments where the antigen is situated. Results showed that the antigen, not like the recombinant expressed in E. coli, located in both cellular fluids and membrane, suggesting that the antigen might not be a skeleton protein as predicted by proteomics.

Content
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Abstract…………………………………………………………………01
Chinese abstract…...……………………………………………………02
Chapter 1: introduction...……………………………………………….03
Chapter 2: materials and methods……………...……………………....10
Chapter 3: results…………………………………………………….....26
Chapter 4: discussion…………………………………………………...42
Chapter 5: conclusion….……………….…………………….………...47
References………………………………..…………………………….48

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